Monoclonal antibodies to human plasma Protein X al

Authors: Dieter Jenne Ferdinand Hugo Sucharit Bhakdi

Publish Date: 1985/04/01

Volume: 5, Issue: 4, Pages: 343-352

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Abstract

Protein X alias complement Sprotein was isolated by dissociation from purified XCSb9 fluidphase terminal C5b9 complexes with 250 mM deoxycholate and subsequent sucrose density gradient centrifugation and Sephacryl gel chromatography Polyclonal rabbit and monoclonal mouse antibodies were used to preliminarily characterize the protein in human serum and plasma In plasma Protein X yielded a symmetrical immunoprecipitate of α2mobility in a crossed immunoelectrophoresis assay However a second immunoprecipitate of Ohmobility was observed when serum was analysed this precipitate represented Protein X in complex with antithrombinIII The coprecipitation of Protein X with serum antithrombinIII was exploited for establishing a simple screening test for unequivocal identification of monocJonal anti – Protein X antibodies SDSPAGE immunoblotting with monoclonal antibodies showed that Protein X exhibits pronounced microheterogeneity migrating as a diffuse moiety of approx Mr 80–90 000 Additionally a small amount of polymeric aggregates appear to be present in plasma Reduction of disulfide bonds led to liberation of a polypeptide of approx 15 K as discerned by twodimensional SDSPAGE immunoblotting Protein X is not cleaved to lower molecular weight entities during the process of blood coagulation or during formation of fluidphase terminal complement complexes The plasma concentrations in healthy adults were in the range of 500–700 pg/ml The availability of methods for isolating Protein X and raising monoclonal antibodies will facilitate further studies on the dual role of this protein in the terminal complement and coagulation cascades

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